Because a radiopharmaceutical consists of a radionuclide and a biochemical, two considerations apply in designing or developing a radiopharmaceutical, one relating to the radionuclide and the other relating to the biochemical.

Selection of a Radionuclide. The choice of a radionuclide for imaging purposes is chiefly dictated by the necessity of minimizing the radiation dose to the patient and the detection characteristics of present-day nuclear medicine instrumentation.

To minimize the radiation dose to the patient, a radionuclide should have as short a half-life as is compatible with the biologic phenomena under study. For example, a radionuclide with a 1-hour half-life, despite its smaller radiation dose, cannot be used in studies of physiologic or metabolic functions that span months. A suggested rule of thumb in this connection is that the physical half-life of the radionuclide should be about 0.693 × T_obs, where T_obs is the time interval between the time of administration of the radionuclide and the time at which measurement or imaging is to be performed. Other requirements of a radionuclide depend on its intended use, either with a scintillation camera (single-photon detection, Chapters 10 and 14) or with positron emission tomography (coincidence detection, Chapter 15), as we outline next.

For Use with Scintillation Camera and Single-Photon Emission Computed Tomography (SPECT) Imaging. It should not emit any corpuscular radiations (e.g., β particles, conversion electrons), because these result in a radiation dose to a patient without providing any benefit. A radionuclide should preferably emit a monochromatic (single-energy) γ-ray with energy between 100 and 300 keV. The upper limit of the desired energy of the γ-ray is the consequence of the detection characteristics of the scintillation camera. As the energy of γ-rays increases, they become more and more penetrating; therefore, a smaller number of them interact within the detector. This reduces the sensitivity of the system. The concept of sensitivity and its importance in nuclear medicine is discussed in later chapters. The lower limit of the desired energy of the γ-ray
is arrived at from the consideration of attenuation of γ-rays in the patient. Because γ-rays should be able to penetrate the patient’s body effectively, their energy has to be high enough to be transmitted out of the patient’s body; hence, the lower limit (Chapter 6).

In addition, a radionuclide should be available easily, economically, and in an uncontaminated form. Technetium-99m, with its 6-hour half-life and 140-keV γ-ray emission, little emission of corpuscular radiation, and easy economical availability from a generator, comes very close to fulfilling the above requirements. This accounts for its wide use in nuclear medicine.

For Use with PET Imaging. The ideal radionuclide for positron imaging is one that does not emit γ-rays other than 511 keV rays generated following positron emission and annihilation, emits positrons with small average energy (minimizing positron range; see Chapters 6 and 15), and is, hopefully, produced by a generator. In addition, it should provide clinically or physiologically important information that is not readily available with single-photon imaging.

Selection of a Chemical. Besides being nontoxic in the desired amounts, the choice of the biochemical or pharmaceutical substance in a radiopharmaceutical is dictated by the requirement that it be distributed or localized in the desired organ or compartment and that the uptake by that organ (or part of the organ) in a normal condition differs substantially from uptake in a pathologic condition. This is generally expressed as target to nontarget ratio. The higher the ratio, the higher the contrast in the image and easier it becomes to visualize a disease (see Chapter 16).

To help in the selection of a suitable biochemical, a wealth of information has been acquired in the field of pharmacology. A number of physiochemical variants determine or affect the distribution and localization of drugs in tissues. Three important determinants in this regard are route of administration, blood flow to the organ or tissues, and extraction by the tissues. Radiopharmaceuticals, with few exceptions, are nearly always administered intravenously, primarily because this is the fastest way to introduce a drug into the circulatory system of the body. Blood flow or perfusion (which can be severely affected in diseases) essentially determines the fraction of the administered dose that will be delivered to a particular organ or tissue during the first transit (10 to 20 seconds). Because blood serves as a carrier for the drug, another property, binding to plasma proteins, plays an important role in the localization of a drug or a chemical in a given tissue. In general, drugs or chemicals strongly bound to plasma proteins remain in blood for a longer period of time (hours to days) and localize to a lesser extent in tissues than those not so tightly bound to plasma proteins.

Extraction of a drug or chemical from circulation and localization in tissue may occur in a number of ways. Simple diffusion, filtration through small pores in the membranes, active transport, receptor binding, and phagocytosis are some examples. As can be seen from Table 5.1, all of these mechanisms are used in the development of radiopharmaceuticals.

When appropriate radionuclides and chemicals have been selected, the following steps are involved in the eventual development of a radiopharmaceutical.

Chemical Studies. These are aimed at establishing the best method of radiolabeling the chemical, defining the optimum condition of labeling and in vitro stability, and determining the nature and the extent of the radiochemical impurities.

Animal Distribution and Toxicity Studies. The main purpose of these studies is to determine bio-distribution of labeled material and establish safe amounts (radioactivity mainly, because the mass of the chemical used is in trace amounts) of the radiochemical that can be administered to humans without subjecting them to undue risk. Bio-distribution establishes the pattern of distribution (major organ or tissues of uptake) of the radioactivity at different times after the administration of the radiochemical in animals considered normal (control) and those in which the appropriate pathologic condition has been induced. From these, one estimates the optimum time for imaging after administration of the radiopharmaceutical and radiation dose delivered to various tissues.

Human or Clinical Studies. Because bio-distribution of a radiopharmaceutical in animals may be different from that in humans, initial studies (phase I) in only a small number of humans are performed to establish the distribution patterns, clearance time, mode of excretion, and optimum imaging time for the radiopharmaceutical. In phase II, these studies are extended to include patients with known diseases and provide further evidence of safety and initial proof of the diagnostic or therapeutic efficacy and final estimates of radiation doses to various human tissues. Finally (phase III), a large series of patients are studied that establishes the overall usefulness (i.e., safety and efficacy) of the agent.

Human or clinical studies are performed as an Investigational New Drug under a Notice of Claimed Exemption to the FDA. Once these data are collected, a New Drug Application is submitted to the FDA, which must approve it before its commercial use.

Because all radiopharmaceuticals are intended eventually for human use, strict quality control is very important. To ensure optimum quality, the following properties of a radiopharmaceutical must be considered.

Radionuclidic Purity. Ideally, the radiopharmaceutical should contain only the desired radionuclide. Often, however, it is not possible to avoid some contamination by other radionuclides, and therefore it is essential to hold this contamination to a low level. A contaminating radionuclide does not add to the diagnostic information, but it does increase the radiation dose to the patient and, in many cases, may degrade the image quality. A good example is provided by the radionuclide ¹²³I, which is difficult to produce without ¹²⁴I as radio-contaminant. Iodine-124, besides significantly increasing the radiation dose to the patient, degrades image quality because of its emission of high-energy γ-rays.

The amount of impurity is generally given as µCi (KBq) of radio-contaminant per µCi (KBq) or mCi (MBq) of the desired radionuclide. Sometimes the limit of the allowable contamination is set by governmental agencies, as in the case of ^99mTc, for which the amount of ⁹⁹Mo must not exceed 0.15 µCi for each mCi of ^99mTc. In cases where no such limits are prescribed, the rule of thumb is to keep the radiation dose to the patient from the radio-contaminants to <10% of that due to the radionuclide of interest.

Another important aspect of radionuclidic purity is that it does not stay constant with time. Where the half-life of the desired radionuclide is shorter than that of the radio-contaminant, radionuclidic purity degrades with time and vice versa. For example, because the half-life of ¹²⁴I, a common radio-contaminant in ¹²³I, is longer than ¹²³I, the radionuclidic purity is best at the time of
the production of this radionuclide, and as the radionuclide is stored, it becomes progressively less pure.

The most common method of determining the nature and extent of radionuclidic impurity is with γ spectroscopy using a NaI (Tl) or Ge (Li) detector, both of which are discussed in Chapter 8.

Radiochemical Purity. Because a radionuclide may form several compounds with a given chemical, it is important to ascertain that a given radiopharmaceutical is in the desired chemical form. Any radiochemical impurities present should be precisely stated. In this regard, it is also important to consider that although a radiochemical may be pure to begin with, it may not be stable over a period of time as a result of the action of radiation or the nature of the chemical itself. To avoid this deterioration, the radiochemical should be stored properly according to the instructions of the manufacturer. For example, radioiodinated human serum albumin (RIHSA), which, among other things, is used as a blood pool scanning agent, may be 99.9% pure when freshly prepared. With time, however, some of the radioiodine becomes free. The amount of free radioiodine strongly depends on storage conditions. The contamination with free radioiodine is several times higher if RIHSA is stored at room temperature than if it is refrigerated. A significant amount of free radioiodine will interfere with the intended study.

A common method for the detection of radiochemical impurities is thin-layer or paper chromatography.

Chemical Purity. A radiopharmaceutical should contain only the desired chemical. In the final preparation of the radiopharmaceutical, there may be a number of chemicals involved besides the radiochemical of interest. These chemicals should be compatible with each other in vitro and safe for the patient. In addition, these must not distort the in vivo function of the main chemical. Aluminum breakthrough in a ⁹⁹Mo-^99mTc generator is an example of potential chemical impurity.

Sterility. A radiopharmaceutical should be sterile (i.e., free from any microbial contamination) and therefore should be tested to this effect before use in patients. In the case of radiopharmaceuticals labeled with short-lived radionuclides (^99mTc and ^113mIn), where prior testing of the final product is not feasible, the sterility of the labeling technique should be tested adequately and periodically.

Apyrogenicity. Even if the preparation is sterile, it may still contain pyrogens that, when intravenously administered to a patient, may cause a reaction. A radiopharmaceutical, therefore, should also be tested for pyrogenicity before use in humans. If this is not feasible, as in the case of short-lived radionuclides, the apyrogenicity of the technique should be ascertained properly and periodically.

Because of the very attractive physical characteristics of ^99mTc, a variety of chemicals have been labeled with this radionuclide, even though the exact mechanism of technetium labeling with these compounds is often not known. Technetium-99m, in the form of sodium pertechnetate (Na^99mTcO₄), is easily obtained in the laboratory from a ^99mMo-^99mTc generator. The labeling of most chemicals by ^99mTc is achieved by first reducing the pertechnetate to ionic technetium (mostly Tc4+) and then complexing it with the desired chemical. The common agent used for reducing purposes is stannous chloride (SnC1₂). Because the half-life of ^99mTc is short (6 hours), most labeling has to be performed “in-house.” This is greatly simplified by the use of sterile and pyrogen-free kits, in which all the desired chemicals are premixed and held together in a lyophilized state under an inert atmosphere (nitrogen gas), except the radionuclide (hence, quite often referred as “cold kit”). To label a particular chemical compound, it is necessary only to introduce a known amount of sterile and pyrogen-free sodium ^99mTc pertechnetate into the kit vial; the labeled compound is ready to use within a few minutes.

Three parameters—labeling efficiency, in vitro stability, and in vivo stability—are important considerations in the selection of a kit. Labeling efficiency is defined as the percentage of total radioactivity present in the kit that is tagged to the appropriate molecule or compound. For most
kits currently in use in nuclear medicine, labeling efficiencies under optimum conditions are in excess of 90%, sometimes even reaching as high as 99%. The remainder of the radioactivity (which is not tagged to the desired compound) is present as radiochemical impurity. In kits that use SnC1₂ as the reducing agent, radiochemical impurities are, in general, of two forms: free pertechnetate (which was not reduced) and reduced or hydrolyzed technetium (which was reduced but did not tag to the compound of interest). Reduced or hydrolyzed technetium sometimes forms a colloid with excess tin present in the kit. This is also a radiochemical impurity but in a different form.

The in vitro stability of a labeled compound determines the time it can be stored on the shelf without significant deterioration. A high in vitro stability allows the compound to be labeled once and then be used in a number of patients at different times of the day. Both labeling efficiency and in vitro stability of compounds prepared from kits can be maximized by observing a few simple precautions, such as using only oxidant-free sodium pertechnetate solution and ensuring no air (oxygen in air, being an oxidizer, reacts with the reducing agent and thus competes with pertechnetate for the reducing agent) is introduced into the reaction vial during the labeling procedure. The in vitro stability of a kit can also be extended by the use of preservatives in the reaction vial, as is done by some manufacturers, and/or by storing the labeled material at low temperatures. We should caution here that in vitro stability of a labeled compound is different from in vitro stability of the cold kit or the chemical compound itself.

The in vivo stability of a labeled compound determines how closely the distribution of the radiolabeled compound in the biologic system parallels that of the unlabeled compound. The distribution of a labeled compound should be similar to the unlabeled compound at least for the duration of the study.

The distribution and use of the most common technetium-labeled compounds in nuclear medicine are described below. Most of these compounds can be easily and rapidly prepared from the commercially available kits.

Technetium-99m Pertechnetate (^99mTc). This radiopharmaceutical is obtained directly from the ⁹⁹Mo-^99mTc generator using saline as the eluting solution. In biologic systems, it behaves similarly to iodine. After oral or intravenous administration, it is selectively concentrated in the thyroid, salivary glands, stomach, and choroid plexus. Disappearance of the pertechnetate from plasma is a multiexponential function. About 50% of the compound is rapidly diluted into extravascular spaces within 15 to 20 minutes. The remaining amount disappears from the plasma with a half-life of about 3 hours. About 20% to 30% of the injected dose is excreted eventually in feces at a slow rate.

The stomach, which is the major organ of uptake, contains 20% to 25% of the injected dosage at 4 hours. So much radioactivity remains in the stomach even after 24 hours that it is not advisable to perform imaging of an abdominal organ using ^99mTc-labeled radiopharmaceuticals (or whose principal γ emission is 170 keV or below) on a patient who had imaging performed with ^99mTc pertechnetate up to 48 hours before the intended study.

Technetium-99m pertechnetate is presently used for thyroid, salivary gland, and stomach imaging. It is no longer used as a brain imaging agent.

Technetium-99m-Labeled Sulfur Colloid. This radiopharmaceutical is easily prepared using commercially available kits. Colloids in general are removed from the bloodstream by the reticuloendothelial (RE) cells of the body. The relative distribution of the colloids among the RE cells of the various organs depends on factors such as the size, nature, and amount of colloidal particles; blood supply to the organ; and on other physiologic and pathophysiologic considerations. In the case of colloidal sulfur tagged with technetium-99m (particle size ˜0.3 µm), about 70% to 80% of the injected dosage is localized in the liver within 10 to 20 minutes of intravenous administration. Of the remaining amount, about 3% is deposited in the spleen and about 15% to 20% is localized in the bone marrow. This agent is, therefore, primarily used for liver, spleen, and bone marrow imaging.

Another ^99mTc-labeled compound sometimes used for liver, spleen, and bone marrow imaging is albumin microaggregates (particle size ˜1 µm).

Technetium-99m-Labeled Macroaggregated Albumin (MAA, Macrotec, or Technescan). This radiopharmaceutical is primarily used in lung imaging. Within a few seconds after intravenous administration of ^99mTc MAA, 90% to 95% of the injected dosage is trapped in the capillary and precapillary bed of the lungs. For effective lung localization, the albumin macroaggregates must be between 15 and 75 µm in size. The biologic half-life of ^99mTc MAA in the lungs is about 8 to 12 hours. The ^99mTc albumin macroaggregates are broken down into smaller (micro) particles that are then taken up by the RE cells of liver and spleen.

Technetium-99m-Labeled Pyrophosphate (PYP), Methyl Diphosphonate (MDP) and Oxidronate (HDP). These radiopharmaceuticals are primarily used for bone imaging. After an intravenous injection, about 50% to 60% of the injected dosage is localized in the skeleton within 15 to 20 minutes. The remainder of the dose is distributed in soft tissue and plasma, from which it is excreted slowly in the urine. About 20% to 30% of the injected dose is either excreted or taken up by the kidneys within 3 hours of the injection.

The use of ^99mTc PYP and ^99mTc MDP for the detection of myocardial infarctions is now well established. ^99mTc PYP is also now used for imaging of cardiac amyloidosis.

Technetium-99m-Labeled Human Serum Albumin. This radiopharmaceutical is primarily used for blood pool imaging such as heart or placenta. After intravenous administration, it is retained in the plasma for a long period of time. However, ^99mTc-labeled albumin is not as stable in vivo as albumin labeled with radioiodine or radio-chromium. Therefore, it is not the preferred agent for plasma volume determination.

Technetium-99m-Labeled Red Cells. These are primarily used for blood pool imaging, detection of sites of gastrointestinal bleeding and cardiac first pass, and are preferred over ^99mTc-labled human serum albumin. Labeling of red cells with radionuclides, in general, is a complex and time-consuming procedure. Simpler methods were developed to label red cells in vivo with ^99mTc. This increased the popularity of ^99mTc-labeled red cells over that of ^99mTc-labeled albumin for blood pool scanning, particularly for the heart. In this method, two steps are involved. In step 1, a patient’s red cells are coated with stannous ion (Sn²⁺). This is achieved simply by injecting about one-fifth of a “cold” PYP kit into the patient. A cold kit is a preparation reconstituted using saline only (i.e., without any radioactivity). In step 2, performed about 30 minutes later, the desired amount of the ^99mTc radioactivity in the form of pertechnetate is administered to the patient as a second injection. The “pretinned” red cells in the patient quickly reduce the pertechnetate to ionic technetium that then readily binds to the red cell surface. Little free pertechnetate or reduced technetium remains in circulation. Most radioactivity (90%) is bound to the patient’s red cells.

The second step—tagging of ^99mTc to the pretinned red cells—can also be performed in vitro by withdrawing pretinned red cells from the patient and incubating them with the desired amount of ^99mTcO^–₄. The red cells thus labeled may be heat damaged, if desired, by heating them in a water bath at 50°C for half an hour. Damaged red cells are used to image the spleen, without interference from the liver, which is the case when sulfur or other colloids are used to image the spleen. A kit, UltraTag RBC, is also available commercially for labeling red blood cells in vitro. In vitro labeling is the most common method now. The in vivo method described before does not result in as efficient a labeling for gastrointestinal bleeding studies, so it is utilized less.

Technetium-99m-Labeled 2,3-Dimercaptosuccinic Acid (DMSA). This radiopharmaceutical is the agent of choice when morphology of the renal cortex is of interest. After an intravenous administration, this radiopharmaceutical is quickly mixed with the plasma volume, from where it is cleared with a half time of about 1 hour. By 2 hours, between 40% and 50% of the injected dosage is taken up by the renal cortex, and about 15% is excreted in urine. Because of rapid in vitro decomposition of ^99mTc-labeled DMSA, it should be stored in a refrigerator and used within half an hour of labeling.

Technetium-99m-Labeled Diethylenetriamine Pentaacetic Acid (DTPA, Pentetate or Techniplex). This radiopharmaceutical is used
primarily for kidney imaging. After intravenous administration, ^99mTc DTPA is rapidly cleared by the kidneys. The biologic half-life in plasma of DTPA chelates in humans is about 15 minutes. Over 80% of the injected dosage can be recovered in urine between 2 and 3 hours postinjection.